We propose to isolate purified adenylate cyclase, the hormone sensitive enzyme that catalyzed the conversion of ATP to cyclic AMP, from rat liver plasma membranes solubilized with Lubrol-PX, a nonionic detergent. This task can be accomplished by utilizing anion exchange chromatography, gel filtration and ligand-specific affinity chromatography on Sepharose-GTP. We will study the effects of fluoride and guanyl nucleotides on purified adenylate cyclase in the absence of contaminating nucleotide phosphohydrolases. Increased enzyme activity effected by guanyl nucleotide interactions with allosteric sites on the enzyme will be related to protein conformational changes, as measured by spectroscopic techniques (circular dichroism and spectrofluorescence). In addition, the possibility will be explored that the glucagon receptor is associated with the solubilized enzyme but that it is functionally uncoupled; we will attempt to restore hormone responsiveness to the adenylate cyclase by modifying the solvent milieu through substitution of detergents and phospholipids. Should glucagon sensitivity be restorable, we will examine hormone induced alterations in conformation of this enzyme-receptor complex and relate these findings to direct assay of adenylate cyclase activity as a consequence of glucagon action on the soluble hormone-resensitized enzyme.